The goal of this proposal is to improve systemic therapies of colon cancer using self-assembled nanomaterials that can deliver potent anticancer miRNA and then degrade in cancer cells to active small molecule modulators of dysregulated polyamine metabolism. Despite tremendous therapeutic potential, clinical translation of miRNA faces major unsolved pharmaceutical delivery challenges. Due to the involvement of multiple mutations in tumorigenesis and tumor progression, combination of miRNAs with modulators of polyamine metabolism has significant therapeutic potential. The fact that polyamine metabolism is downstream from many oncogenes and tumor suppressor pathways make it a logical target for such combination miRNA therapy approaches. Our objective is to develop polyamine prodrugs (PaPs) that can modulate dysregulated polyamine metabolism and encapsulate and systemically deliver anticancer miR-34a. The hypothesis is that self-immolative PaPs based on modulators of polyamine metabolism will deliver miR-34a to the tumors, which will result in enhanced combination effect due to the downregulation of tumor polyamine biosynthesis and upregulation of polyamine catabolism and restoration of important cell growth and death-regulatory functions due to miR-34a. We will accomplish the objectives in three specific aims: (1) we will optimize formulation of tumor-penetrating PaP/miR-34a nanoparticles that deliver miRNA and modulate polyamine metabolism. Based on encouraging anticancer in vivo activity in our preliminary studies, we hypothesize that particle modification with tumor-penetrating iRGD peptide and with stabilizing superhydrophobic fluorinated moieties will result in efficient systemic delivery. (2) we will determine the mechanism of action of PaP/miR-34a nanoparticles in vitro. Our results indicate that PaPs are effective in reducing tumor cell growth and induction of apoptosis, but the precise mechanism of action of the nanoparticles and how it relates to their intracellular trafficking, disassembly and rate of polyamine analog release is unknown. We will ascertain the mechanisms of action and determine which composition strategies are most effective in producing strong antitumor effect. (3) we will test the in vivo efficacy of the particles in colon cancer using human tumor xenografts and syngeneic immune competent tumor models. We will conduct comprehensive evaluation of anticancer activity and survival advantage of PaP/miR-34a in models relevant for human disease. Contribution of the antitumor immunogenicity to the efficacy will be also studied due to known effects of polyamine analogs on increasing the antitumor immune response. We predict that we will be able to prepare nanoparticles with improved anticancer activity and prolonged survival. The proposed integrative approach is innovative because of the dual-function design of the PaP polymers as modulators of polyamine metabolism and miRNA carriers. The research is significant because it will address major barriers in developing drug/nucleic acid nanotechnology for systemic treatment of cancer and establish a widely applicable and versatile platform for combination delivery systems that target polyamine metabolism as a way of improving anticancer therapies.